One of the leading causes of morbidity and mortality in Western societies is atherosclerosis. NADPH oxidase contributes to atherosclerosis through the indirect activation of macrophages leading to the internalization of oxidized low density lipoproteins (LDL). Chronic inflammation in activated macrophages contributes to atherosclerosis, while increased production of oxygen/nitrogen radicals leads to chemical modifications of extracellular LDL. Because macrophages are positioned at the cross-roads of lipid metabolism in vessel walls, they are important cells in the cellular pathology of atherosclerosis. Components of the endogenous cannabinoid (eCB) system are vital to atherosclerosis development, since the CB system has been found to play an important role in the amelioration of atherosclerosis. The eCB system has several components, including the G-protein- coupled cannabinoid receptors (CB1 and CB2); their endogenous endocannabinoid ligands, 2- arachidonoylglycerol (2-AG) and anandamide (AEA); and biosynthetic enzymes that produce these compounds and hydrolytic enzymes that degrade them. For instance, CB2 signaling has been shown to upregulate immunoprotective and anti-oxidative pathways, whereas CB1 signaling has opposite effects. We hypothesize a mechanistic link between CD36 activation and NADPH oxidase activity, which will lead to enhanced biosynthesis of 2-arachidonoylglycerol (2-AG) via a signaling pathway that activates diacylglycerol lipase beta (DAGL). Activation of CB2-mediated signaling by the resulting enhanced endocannabinoid tone will potentially reduce oxidative stress in macrophages. The released 2-AG is subsequently catabolized by surrounding vessel wall cells. We believe this link will lead to enhanced biosynthesis of 2-AG via a signaling pathway that activates diacylglycerol lipase. We intend to first prove that ligation of macrophage CD36 by oxLDL will activate NADPH oxidase and increase ROS production. We will use both human macrophages and mouse macrophages derived from Nox 2-deficient mice to demonstrate cause and effect. Secondly, we will demonstrate that increased levels of superoxide causes enhanced 2-AG biosynthesis within the macrophage, and that upregulation in eCB production is an adaptive response to oxidative stress. Finally, we will examine the catabolism of 2-AG by cultured endothelial and smooth muscle cells and profile the serine hydrolases that are present in these cell types using an activity-based protein profiling (ABPP)-MudPIT approach that our laboratory has previously done using human macrophages. We expect that enhancing endocannabinoid tone within the vessel wall may be a valuable strategy to reduce the occurrence of atherosclerosis.